Abstract The overall goal of this application is to characterize newly discovered ultra-small parasitic bacteria that have extremely reduced genomes and show increased abundance in gingivitis and periodontitis. In the previous funding cycle, we successfully cultivated and sequenced TM7x, the first member of the uncultivated TM7 phylum from humans. Strain TM7x is unique among all bacteria, it has an ultrasmall size (200-300 nm) and lives on the surface of a host bacterium, a relationship that had never been reported in the human microbiome or in the Bacteria domain. With a highly-reduced genome, TM7x cannot synthesize any of its own amino acids, vitamins or cell wall precursors and must parasitize other oral bacteria which impacts their growth. Of particular note is that TM7 belongs to the Candidate Phyla Radiation (CPR), a recently discovered subdivision in the domain Bacteria that comprises >26% of the known bacterial diversity with an estimated 70 uncultivated Phyla with reduced genomes. To date, TM7x is still the only reported cultivated representative of the entire CPR, putting our team in a unique position to make significant advancements in the field and facilitate fundamental discoveries on these ultra-small human associated bacteria, as well as the CPR group as a whole. Our preliminary data revealed that the relationship of TM7x and its bacterial host, an oral Actinomyces odontolyticus strain XH001 is a highly-regulated symbiotic interaction in which TM7x displays both symbiotic phase (co-existing with host) and virulent parasitic phase (inducing host cell death). This intriguing relationship is similar to the one observed for temperate bacteriophages and their hosts where phages are capable of switching between lysogenic and lytic cycle, and entails physiological and ecological consequences. In this application, we aim to achieve the following two goals. 1) To identify the molecular components and regulatory pathways governing the relationship between TM7x and its host; 2) To investigate the range of bacteria that TM7x interacts with. In particular, the mechanism of host killing encoded within a bacterial parasite with a reduced genome is both fundamentally novel and clinically relevant. The ability to infect and kill multiple bacterial hosts may allow TM7x to influence the oral community dynamics and structure, thus modulating community and impacting microbiome during health and diseases. The success of the study would greatly advance the developing research field of these ultra-small bacteria, expand our knowledge on this novel microbial symbiosis found in humans, as well as the potential impact of CPR organisms on oral microbiome ecology.